The present invention relates to treatment and prevention of Human Immunodeficiency Virus (HIV) by using a live attenuated nef deleted HIV-1 virus vaccine.
Human Immunodeficiency Virus (HIV) is the primary etiologic agent for the acquired immunodeficiency syndrome (AIDS). HIV exhibits high genetic variation, which results in a wide variety of biological phenotypes displayed by various strains of the virus and also by the same strain of the virus in a single patient at different times. Such phenotypic heterogeneity is exhibited in replication kinetics, susceptibility to serum neutralization, anti-viral drug resistance, induction of cytopathicity and host-cell range specificity. The two main of the Human Immunodeficiency Virus, subtypes HIV-1 and HIV-2, are members of a group of closely related human and non-human primate lentiviruses, which are RNA retroviruses.
Infection in humans by HIV leads to progressive deterioration of cell mediated immune system making the victim susceptible to a variety of opportunistic infections, such as pneumocystis carinii pneumonia (PCP) and tumors such as Kaposi's sarcoma (KS). It is known that the mechanism of the destruction of the immune system, centers on the cytopathic effect of HIV on CD4+ THELPER lymphocytes which are instrumental in proper functioning of cell mediated immunity.
AIDS and HIV human infection initially involved homosexual men, intravenous drug users, and hemophiliacs in the United States and Europe. However, heterosexual infection has become common and rampant in Africa (particularly in Rwanda, Burundi, Zaire and Kenya), Brazil, India, Myanmar and Thailand. According to the World Health Organization, in excess of 40,000,000 people worldwide are estimated to be infected with the HIV. The available data indicates that almost all of these HIV infected individuals will die for lack of an effective treatment.
Humoral antibody response mediated by B Lymphocytes is usually strong in infected individuals with high antibody titers, especially those infected at the envelope proteins gp120, gp41 and gag proteins p24, p17 and p15. Unfortunately, high humoral antibody response in humans does not provide any protection from continued and relentless infection and progression of the HIV disease. This, result is mainly due to cell to cell transmission of infection and inhibition of cytotoxic T lymphocytes, perhaps by inhibition of the IL-2 (Interleukin-2) signaling. The US National Institute of Health recently abandoned phase III and phase IV trials of vaccines derived from various viral proteins of HIV because of disappointing results in earlier phases. Similarly, cellular response against HIV is' initially strong with an increase in cytotoxic (“killer”) T lymphocytes (CTL). Unfortunately, this response breaks down soon after infection due to genetic variations in the gag CTL epitopes which allows the virus to escape CTL recognition. (Phillips R E et al; Nature 345:453, 1991).
Various drugs have also been approved for treatment of HIV infection such as zidovudine which interfere with the virus's nucleotide sequencing. While these were felt to be very promising in the earlier stages, development of resistance to them has caused a considerable amount of disappointment and frustration.
A variety of other approaches have been postulated. Professor Jonas Salk, in his commentary in the publication Nature noted that as the disease progresses, titers of antibodies to gp41 and virus neutralizing antibodies remain constant, but the level of anti-p24 antibodies, which correlates with the presence of antibody dependent cell cytotoxicity (ADCC) and antibody to reverse transcriptase, decline. He proposed treatment of symptomatic HIV infected patients with sera from asymptomatic HIV infected patients. He further hypothesized that HIV immunogens given to HIV infected patients would be protective. (Salk J; Prospects for the control of AIDS by immunizing seropositive individuals. Nature 327:473-476, 1987).
Live-attenuated viruses and dead virions have been hypothesized but no researcher has yet tried these either for prevention or treatment of HIV infection in humans in a meaningful manner.
SIV (Simian Immunodeficiency Virus) is a primate lentivirus with various strains that affect African green monkeys, macaque monkeys, sooty-mangabee monkeys, rhesus monkeys and chimpanzees. SIV infection in non-human primates is widely used to study the physiology and pathology of the SIV primate lentiviruses. A great deal of research has been done by attempting to infect monkeys with artificially created mutants of the SIV to determine their relative infectivity. Many of these studies focused on the role of the nef gene in the physiology of virus life cycle. The nef gene is present in all primate lentiviruses sequenced to-date. The gene consists of an open reading frame beginning within or immediately after the 3′ end of the env gene and overlaps the U3 portion of the 3′ long terminal repeat. The gene was previously named F, 3′-orf or B-orf. It is expressed in vivo as determined by antibodies to the nef gene product in infected individuals. Luria et al have shown that at least some nef gene products block the induction of IL-2 (Interleukin-2) mRNA in lymphoid cells triggered by activating agents PMA, PHA and/or antibodies against CD3, TCR or CD2 (Luria S, Chambers I, Berg P; Proc Natl Acad Sci USA 88:5326, 1991). Kestler et al have found rapid reversion of stop codon point mutations in nef to open forms, in vivo, demonstrating selective pressure for open, presumably functional, forms of nef. (Kestler H W et al; Cell, 65:651, 1991). It was further shown that nef is necessary for vigorous SIV virus replication in rhesus monkeys, for maintaining normal virus loads, and for induction of the SIV disease. Animals inoculated with nef-deletion mutants have remained disease free for at least 3 years, while wild-type virus infected animals all developed AIDS and died. It has also been demonstrated that nef deletion increases viral replication but it is postulated that the responses to nef deletion are different in vivo and in vitro. (Gibbs J S and Desrosiers R C in Human Retroviruses, Cullen B R, ed, Oxford University Press, NY, 1993).
Derosiers, R C (WO 92/00987 as well as U.S. Pat. No. 5,851,813) teaches a nef impaired construct in various primate lentiviruses. Desrosiers disclosed only a “non-revertible null mutation” in the nef gene and not a removal of substantially all of the open reading frame (ORF) from the nef gene. It was later demonstrated that even the “non revertible” null mutations recombine to the wild-type viruses. (Ref: Alexander L, Illiyinski P O, Lang S M, Desrosiers R C et al: Determinants of Increased Replicative Capacity of Serially Passaged Simian Immunodeficiency Virus with nef Deleted in Rhesus monkeys Journal of Virology 77:12, June 2003, 6823-6835). Desrosiers, suggested the compositions proposed worked as antigenic agents rather than as a signals defect restoration agent. The Desrosiers composition was deemed to have “preventive” benefits when given to healthy subjects, rather than to primates already infected with SIV i.e. as a treatment. In addition the Desrosiers experiments were restricted to chimpanzees and used a completely unrelated virus i.e. SIV, which does not cause disease in humans, just as HIV-1 does not cause disease in chimpanzees. Thus, applying the knowledge that nef deleted SIV imparts immunity in chimpanzees does not translate into null mutations in the nef gene of HIV producing a similar occurrence in humans. Desrosiers never actually injected humans with the compositions proposed. Eventually the Desrosiers proposal proved not to work and caused AIDS in both newborn and adult primates (Refs: Baba T W, Liska V, Khimani A H, Ray N B, Dailey P J, Penninck D, Bronson R Greene M F, McClure H M, Martin L N, Ruprecht R M. Live attenuated, multiple deleted simian immunodeficiency virus causes AIDS in infant and adult macaques. Nat Med (1999 February) 5(2):194-203 and Live attenuated, nef-deleted SIV is pathogenic in most adult macaques after prolonged observation. Hofmann-Lehmann R, Vlasak J, Williams A L, Chenine A L, McClure H M, Anderson D C, O'Neil S, Ruprecht R M AIDS (2003 Jan. 24) 17(2):157-66). This failure was confirmed by Desrosiers. (Alexander L, Illiyinski P O, Lang S M, Desrosiers R C et al: Determinants of Increased Replicative Capacity of Serially Passaged Simian Immunodeficiency Virus with nef Deleted in Rhesus monkeys Journal of Virology 77:12, June 2003, 6823-6835).
It is also known that the nef gene in HIV-1 is well-conserved across various strains and subtypes, any strain may be used to prepare the subject invention. [Kotov A, Zhou J, Flicker P, Aiken C Association of Nef with the human immunodeficiency virus type 1 core. J Virol (1999 October) 73(10):8824-30, Jubier-Maurin V, Saragosti S, Perret J L, Mpoudi E, Esu-Williams E Mulanga C, Liegeois F, Ekwalanga M, Delaporte E, Peeters M. Genetic characterization of the nef gene from human immunodeficiency virus type 1 group M strains representing genetic subtypes A, B, C, E, F, G, and H. AIDS Res Hum Retroviruses (1999 Jan. 1) 15(1):23-32, Shugars D C, Smith M S, Glueck D H, Nantermet P V, Seillier-Moiseiwitsch F, Swanstrom R. Analysis of human immunodeficiency virus type 1 nef gene sequences present in vivo [published erratum appears in J Virol 1994 August; 68(8):5335] J Virol (1993 August) 67(8):4639-50, Wentworth P A, Lee D J, Doe B, Feucht P H, Bathurst I C, Steimer K Characterization of human helper T cell epitopes within HIV-1 nef. Int Conf AIDS (1991 Jun. 16-21) 7(2):26 (abstract no. W.A.20), Cheng H, Hoxie J P, Parks W P. The conserved core of human immunodeficiency virus type 1 Nef is essential for association with Lck and for enhanced viral replication in T-lymphocytes. Virology (1999 Nov. 10) 264(1):5-15]
An HIV treatment which successfully overcomes the problems encountered in the prior art would be a notable advance in the medical arts.